title: Hyperosmolarity-induced up-regulation of claudin-4 mediated by NADPH oxidase-dependent H2O2 production and Sp1/c-Jun cooperation.
abstract: Claudin-4 is exclusively localized in the tight collecting ducts in the renal tubule. We examined what molecular mechanism is involved in the regulation of claudin-4 expression. In Madin-Darby canine kidney cells, hyperosmolarity increased the expression level of claudin-4 and the production of reactive oxygen species, which were inhibited by diphenyleneiodonium (DPI), an NADPH oxidase inhibitor, and manganese (III) tetrakis (4-benzoic acid)porphyrin (MnTBAP), a scavenger of H2O2. Both hyperosmolarity and H2O2 increased p-ERK1/2 and p-JNK, which were inhibited by U0126, a MEK inhibitor, and SP600125, a JNK inhibitor, respectively. Immunoprecipitation assay showed that hyperosmolarity increased the association of nuclear Sp1 with c-Jun, which was inhibited by U0126 and SP600125. In mouse inner medullary collecting duct cells and rat kidney slices, hyperosmolarity increased the expression level of claudin-4, which was inhibited by DPI, MnTBAP, U0126, and SP600125. Hyperosmolarity increased luciferase reporter activity of claudin-4, which was inhibited by U0126, SP600125, Sp1 siRNA, and c-Jun siRNA. The activity was inhibited by the mutation in the Sp1 binding site. Chromatin immunoprecipitation assay and avidin-biotin conjugated DNA assay showed that Sp1 and c-Jun are associated with the Sp1 binding site. These results suggest that hyperosmolarity increases nuclear Sp1/c-Jun complex and the association of the complex with the Sp1 binding site, resulting in the segment-specific expression of claudin-4 in the kidney.
authors.affiliation: Department of Pharmaco-Biochemistry, School of Pharmaceutical Sciences, University of Shizuoka, Shizuoka, Japan. Electronic address: ikari@u-shizuoka-ken.ac.jp.
authors.forename: Akira
authors.initials: A
authors.lastname: Ikari
authors.affiliation: Department of Pharmaco-Biochemistry, School of Pharmaceutical Sciences, University of Shizuoka, Shizuoka, Japan.
authors.forename: Kosuke
authors.initials: K
authors.lastname: Atomi
authors.affiliation: Department of Pharmaco-Biochemistry, School of Pharmaceutical Sciences, University of Shizuoka, Shizuoka, Japan.
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Yamazaki
authors.affiliation: Department of Pharmaceutical Physiology, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama, Japan.
authors.forename: Hideki
authors.initials: H
authors.lastname: Sakai
authors.affiliation: Laboratory of Physiology, School of Food and Nutritional Sciences, University of Shizuoka, Shizuoka, Japan.
authors.forename: Hisayoshi
authors.initials: H
authors.lastname: Hayashi
authors.affiliation: Department of Pharmaco-Biochemistry, School of Pharmaceutical Sciences, University of Shizuoka, Shizuoka, Japan.
authors.forename: Masahiko
authors.initials: M
authors.lastname: Yamaguchi
authors.affiliation: Department of Pharmaco-Biochemistry, School of Pharmaceutical Sciences, University of Shizuoka, Shizuoka, Japan.
authors.forename: Junko
authors.initials: J
authors.lastname: Sugatani
mesh_terms: D000818:Animals; D000873:Anthracenes / Q000494:pharmacology; D001483:Base Sequence; D002070:Butadienes / Q000494:pharmacology; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D062506:Claudin-4 / Q000235:genetics* / Q000378:metabolism; D003513:Cycloheximide / Q000494:pharmacology; D004285:Dogs; D004791:Enzyme Inhibitors / Q000494:pharmacology; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D016166:Free Radical Scavengers / Q000494:pharmacology; D006861:Hydrogen Peroxide / Q000378:metabolism*; D006982:Hypertonic Solutions / Q000494:pharmacology*; D048031:JNK Mitogen-Activated Protein Kinases / Q000378:metabolism; D007685:Kidney Tubules, Collecting / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D061985:Madin Darby Canine Kidney Cells; D008954:Models, Biological; D008969:Molecular Sequence Data; D019255:NADPH Oxidases / Q000378:metabolism*; D009570:Nitriles / Q000494:pharmacology; D009994:Osmolar Concentration; D011401:Promoter Regions, Genetic / Q000235:genetics; D016755:Proto-Oncogene Proteins c-jun / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar; D016329:Sp1 Transcription Factor / Q000378:metabolism*; D015854:Up-Regulation / Q000187:drug effects*
chemical_list: D000873:Anthracenes; D002070:Butadienes; D062506:Claudin-4; D004791:Enzyme Inhibitors; D016166:Free Radical Scavengers; D006982:Hypertonic Solutions; D009570:Nitriles; D016755:Proto-Oncogene Proteins c-jun; D016329:Sp1 Transcription Factor; C113580:U 0126; C432165:pyrazolanthrone; D003513:Cycloheximide; D006861:Hydrogen Peroxide; D019255:NADPH Oxidases; D048049:Extracellular Signal-Regulated MAP Kinases; D048031:JNK Mitogen-Activated Protein Kinases
keywords: ABCD; ChIP; Claudin-4; ERK; GSH; H(2)O(2); Hyperosmolarity; JNK; MAPK; MDCK; Madin–Darby canine kidney; N-acetyl-l-cysteine; NAC; ROS; Sp1; TJs; avidin–biotin conjugated DNA; c-Jun; c-Jun N-terminal kinase; chromatin immunoprecipitation; extracellular signal-regulated kinase; hydrogen peroxide; mIMCD; mitogen-activated protein kinase; mouse inner medullary collecting duct; reactive oxygen species; reduced glutathione; tight junctions